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0EFFECTS OF SINK OXIDE NANOPARTICLES ON PHOTOPHYSICAL BEHAVIOUR OF CHLOROPHYLL
ELDOST FATALIYEV, ASLAN HEYBATOV
Baku State University,Department of Molecular biology and biotechnology, Baku,Azerbaijan
ISMAT AHMADOV, ZIYADDIN MAMAEDOV Baku State University,Department of Cemical physics of nanomaterials , Baku,Azerbaijan
Abstract. In the presented article, the interaction of zinc oxide nanoparticles with the main photosynthetic pigment chlorophyll was investigated using UV-vis and laser-induced fluorescence spectroscopy. Chlorophyll pigment was isolated from the leaves of 10-day-old bean sprouts by a standard method. Steady-state fluorescence measurements show that zinc oxide quenches chlorophyll fluorescence in a concentration-dependent manner. In the presence of zinc oxide nanoparticles chlorophyll's absorbance decreases, but the concentration increase of nanoparticles zinc oxide does not induce any significant changes. From the recording of fluorescence spectra, it is clear that in bean plants whose seeds are not covered with ZnO nanoparticles, when ZnO nanoparticles are added to the chlorophyll extract and kept in the dark, no drastic change in the intensity of the fluorescence spectrum is observed. However, when ZnO nanoparticles were added to the chlorophyll extract and illuminated, a significant decrease in the intensity of the fluorescence spectrum was observed compared to the control.
Keywords: nanoparticles, bean leaf, chlorophyll, ultraviolet and fluorescence spectra
ВЛИЯНИЕ НАНОЧАСТИЦ ОКСИДА SINK НА ФОТОФИЗИЧЕСКОЕ ПОВЕДЕНИЕ ХЛОРОФИЛЛА
ЭЛДОСТ ФАТАЛИЕВ, АСЛАН ГЕЙБАТОВ, ИСМАТ АХМАДОВ, ЗИЯДДИН
МАМАЕДОВ
Аннотация. В представленной статье взаимодействие наночастиц оксида цинка с основным фотосинтетическим пигментом хлорофиллом исследовалось с помощью УФ-видимой и лазерно-индуцированной флуоресцентной спектроскопии. Пигмент хлорофилл был выделен из листьев 10-дневных ростков фасоли стандартным методом. Измерения стационарной флуоресценции показывают, что оксид цинка тушит флуоресценцию хлорофилла в зависимости от концентрации. В присутствии наночастиц оксида цинка поглощательная способность хлорофилла уменьшается, но увеличение концентрации наночастиц оксида цинка не вызывает существенных изменений. Из регистрации спектров флуоресценции ясно, что у растений фасоли, семена которых не покрыты наночастицами ZnO, при добавлении наночастиц ZnO к экстракту хлорофилла и выдерживании в темноте не наблюдается резкого изменения интенсивности спектра флуоресценции. Однако при добавлении наночастиц ZnO к экстракту хлорофилла и освещении наблюдалось значительное снижение интенсивности спектра флуоресценции по сравнению с контролем._
Introduction
In green plants, the chlorophyll pigment plays an important role in the photosynthesis process as it acts as a light-capturing center. The chemical structure of chlorophyll contains a porphyrin ring with one reduced pyrolle ring with an Mg2+ ion coordinated and a long non-polar phytylic chain that increases the hydrophobicity of the molecule (Fig.1). When chlorophyll is isolated from the chloroplast, the magnesium ion is unstable and can be easily removed by a weak acid (Ssha and Murray, 2018). Therefore various physicochemical parameters that influence the stability of chlorophyll, such as pH of the solution and temperature. Experiments show that chlorophyll is less
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stable compared to the synthetic colorants (Chen X B, et al. 2006). Therefore, any change in the structure and amount of this pigment affects the intensity of the photosynthesis process and ultimately the growth and productivity of the plant. Chlorophyll plays an important role in plants as a photocatalyst and also as an electron donor in the photochemical reactions of photosynthesis. Chlorophyll reduces nicotinamide adenine dinucleotide phosphate (NADP+) by moving photosynthetic electrons released from water hydrolysis in photosystem I (PSI) and photosystem II (PSII) reaction centers (Senge et al. 2014). Chlorophyll has a broad absorption spectrum in the visible wavelength range of light. It has a small band gap, thermal stability, high molar absorption coefficient, highest occupied molecular orbital - lowest unoccupied molecular orbital level energy (Kathiravan et al. 2009b). Therefore, it is easily excited when irradiated with visible light. Chlorophyll and zinc oxide nanoparticles can be of great importance for photoelectric energy conversion processes as semiconductor nanoparticles. Along with chlorophyll pigment, carotenoids also play an important role in protecting plants against biotic and abiotic stress factors as important plant pigments. They protect plants against stresses by quenching harmful singlet oxygen (1O2) species, which are components of reactive oxygen species (ROS) in stressed plants (Ramel et al., 2012). To understand the molecular dynamics of the interaction of biomolecules with nanoparticles, laser-induced fluorescence and UV-vis spectroscopic methods have been successfully applied (Reymond-Laruinaz et al. 2016). These methods allow detection of conformational changes and photophysical parameters of molecules.
ïCH^i-CHjCHj-CHjéH-CHjCHj-CHj-éH-CHj-
Figure 1. Chemical structure of chlorophyll.
In these experiments, the interaction mechanism of chlorophyll with zinc oxide nanoparticles was investigated using steady-state and time-resolved fluorescence spectroscopic and UV-visible spectroscopic methods in both light and dark conditions. Time-resolved fluorescence measurements were also performed to determine the exact quenching mechanism involved in the interaction of zinc oxide nanoparticles with chlorophyll.
Materials and methods
Chlorophylls were extracted from the leaves of Vicia faba (L.), which has been often used as a model system for plant physiological experiments. Bean leaves were collected from the plant grown in a Plant Growth Chamber, cut into small pieces, and added to the acetone in the proportion of 0,1g:10 ml acetone at room temperature. After the bean seeds were covered with ZnO nanoparticles, they were planted in vegetation pots and cultivated in the Plant Growth Chamber phytotron. In the control variant, the seeds were not coated with ZnO nanoparticles. When the seeds germinated and were 10 days old, samples were taken from the second-tier leaves and chlorophyll pigment was separated. Chlorophyll pigment was isolated from both the control variant and the test variant. From both options, 10 ml of acetone solution of chlorophyll pigment was taken and 1 ml of ZnO colloid solution was added. Then the samples were kept in the dark for 7 days in one version and illuminated for 7 days with 9 hours a day in the second version. After 7 days, UV-vis and fluorescence spectra of the samples were recorded.
Optical analyses of the molecular absorption in the 200-800 nm wavelength range were performed in a SPiCORD 250 plus UV-Vis spectrophotometer using a quartz cuvette of 4 mm optical path length at room temperature. The fluorescence measurements were performed using a fluorimeter consisting of a laser excitation source, and a monochromator Cary Eclipse Fluorescence Spectrometer (Varian). The fluorescence spectra of the samples were obtained at wavelengths between 450 and 800 nm with excitation at 457 nm. All measurements were carried out at room temperature using a quartz cuvette with four polished faces and 10 mm optical path length.
Results and discussion Characterization of the nanoparticles
In figure 2 gives the X-ray diffraction pattern (A) and UV-vis spectra of zinc oxide nanoparticles. in the X-ray diffraction pattern have been determined the position, width, intensity and full width at half maximum of the peaks. Line broadening observed in the X-ray diffraction peaks is indicative of the nano-scale dimension of the zinc oxide nanoparticles. The X-ray diffraction pattern of zinc oxide nanoparticles shows sharp peaks at 31.77, 34.44, 36.30, 47.50,56.60, 62.91, 68.08 and 69.09 that have been assigned to the 100, 002, 101, 102, 110,103, 200, 112 and 201 lattice planes respectively. These lattice planes have been assigned to the hexagonal wurtzite phase of zinc oxide (Khalil et al.2014). in figure 2B shows the UV-vis spectra of zinc oxide nanoparticles. in UV-vis spectra of zinc oxide nanoparticles the absorbance maximum peak is 273 nm.
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Figure 2. X-ray diffraction pattern (A) and UV-vis spectra of the ZnO nanoparticles solution
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UV-vis spectroscopy study
Zinc interaction with chlorophyll was examined earlier and has been shown that Zn rather forms "chelate" complexes with chlorophyll. In Zn incubated pigment solutions 5 days after incubation, just a very small hypsochromic effect of the Qy band in UV-vis spectra was observed (Zvezdanovi et al.,2007), then "central" complexes, providing a shifted equilibrium between "central" and "chelate" Zn-Chl complexes toward the latter ones (Nonomura et al., 1997).
Figure 3. Ultraviolet-visible spectra of 10 ml chlorophyll (Chl) in the presence of 1,0 ml solutions of zinc oxide nanoparticles (ZnO NPs) Figure 3 shows the recorded ultraviolet-visible absorption spectrum of the chlorophyll extract and chlorophyll extract mixed with 1,0 ml zinc oxide nanoparticles. The absorption spectra of chlorophyll show two bands: a blue band at 430 nm and a red band observed at 661 nm . In the presence of zinc oxide nanoparticles chlorophyll's absorbance decreases, but the concentration increase of nanoparticles zinc oxide does not induce any significant changes. The observed decrease in the absorbance value indicates some type of interaction between chlorophyll and zinc oxide nanoparticles which has been further described in terms of the change in fluorescence properties of the sample.
Figure 4 shows the recorded steady-state fluorescence emission spectra of chlorophyll extracts stored in the dark with zinc oxide nanoparticles for 7 days and illuminated for 9 hours each day with excitation at 480 nm, respectively. The blue spectrum in Figure 4 is the spectrum of the chlorophyll extract left in the dark and added 0.1 ml of ZnO nanoparticles. The black spectrum is the spectrum of the chlorophyll extract left in the dark but without ZnO nanoparticles added. The green spectrum is the spectrum of the chlorophyll extract that was illuminated and added 0.1 ml of ZnO nanoparticles. The red spectrum is the spectrum of the chlorophyll extract illuminated but without ZnO nanoparticles added.
Wavelength (nm)
Figure 4. Fluorescence spectra of 10 ml chlorophyll (Chl) in the presence zinc oxide
nanoparticles with 0,1 ml From the recording of fluorescence spectra, it is clear that in bean plants whose seeds are not covered with ZnO nanoparticles, when ZnO nanoparticles are added to the chlorophyll extract and
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kept in the dark, no drastic change in the intensity of the fluorescence spectrum is observed. However, when ZnO nanoparticles were added to the chlorophyll extract and illuminated, a significant decrease in the intensity of the fluorescence spectrum was observed compared to the control. It can be concluded that the interaction between chlorophyll and ZnO nanoparticles is more pronounced under lighting conditions.
Conclusions
The results of the presented research allow us to give an important insight into the photophysical behavior of the interaction between chlorophyll and zinc oxide nanoparticles. Zinc oxide nanoparticles quench the intrinsic fluorescence of chlorophyll through dynamic quenching. The decrease in the value of the binding constant and the negative value of the thermodynamic parameters suggest that van der Waal forces and hydrogen bonding are the main binding forces and that the process is spontaneous and exothermic. In addition, the interaction also involves electron transfer, possibly between chlorophyll and zinc oxide nanoparticles. The results of this study may be useful for understanding the sensitization processes associated with chlorophyll zinc oxide nanoparticles to improve the efficiency of photovoltaic conversions.
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